Expression analysis of taste receptor genes (T1R1, T1R3, and T2R4) in response to bacterial, viral and parasitic infection in rainbow trout, Oncorhynchus mykiss

Highlights

• Three infection models with bacterial, viral and parasitic pathogens were successfully established.

• Significantly expression of taste receptor genes occurred in trout gustatory tissues after infected with pathogens.

• Taste receptor genes may be associated with mucosal immune response in teleost.

Abstract

Emerging evidence suggests that bitter and sweet Taste receptors (TRs) in the airway are important sentinels of innate immunity. TRs are G protein-coupled receptors that trigger downstream signaling cascades in response to activation of specific ligands. Among them, the T1R family consists of three genes: T1R1, T1R2, and T1R3, which function as heterodimers for sweet tastants and umami tastants. While the other TRs family components T2Rs function as bitter tastants. To understand the relationship between TRs and mucosal immunity in teleost, here, we firstly identified and analyzed the molecular characteristics of three TRs (T1R1, T1R3, and T2R4) in rainbow trout (Oncorhynchus mykiss). Secondly, by quantitative real-time PCR (qPCR), we detected the mRNA expression levels of T1R1, T1R3 and T2R4 and found that the three genes could be tested in all detected tissues (pharynx, buccal cavity, tongue, nose, gill, eye, gut, fin, skin) and the expression levels of T1R3 and T2R4 were higher in buccal mucosa (BM) and pharyngeal mucosa (PM) compare to other tissues. It may suggest that T1R3 and T2R4 play important roles in BM and PM. Then, to analyses the changes of expression levels of the three genes in rainbow trout infected with pathogens, we established three infection models Flavobacterium columnare (F. cloumnare), infectious hematopoietic necrosis virus (IHNV) and Ichthyophthirius multifiliis (Ich). Subsequently, by qPCR, we detected the expression profiles of TRs in the gustatory tissues (BM, PM and skin) of rainbow trout after infection with F. cloumnare, IHNV, and Ich, respectively. We found that under three different infection models, the expression of the T1R1, T1R3 and T2R4 showed their own changes in mRNA levels. And the expression levels of the T1R1, T1R3 and T2R4 changed significantly at different time points in response to three infection models, respectively, suggesting that TRs may be associated with mucosal immunity.

Introduction

In vertebrates, gustation is one of the specialized chemosensory system dedicated to the evaluation of food and drink, which has evolved into a dominant regulator and driver of feeding behavior. Gustatory system can detect nutrition-related and harmful compounds in food and trigger innate behaviors to accept or reject potential food sources [1]. In mammals, taste organ (tongue) can be divided into a small palette of qualities containing a lot of taste buds, which functionally perceive sweet, bitter, umami and salty [[2], [3], [4]]. So far, two families of G protein-coupled receptors (GPCRs) for tastants have been identified in taste bud cells-T1Rs for sweet tastants and umami tastants (l-amino acids) and T2Rs for bitter tastants [5], with a large extracellular region composed of two domains: the venus flytrap module (VFTM) and the cysteine-rich domain (CRD) [6]. Generally, the T1R family includes three members: T1R1, T1R2, and T1R3. Among them, T1Rs combine to generate two heteromeric G-protein-coupled receptor complexes: T1R1/3, an umami sensor, and T1R2/3, a sweet receptor [7]. T2Rs consist of approximately 30 members [8]. In contrast, T2R4 protein has a single structural feature [6]. In teleost, the taste buds are widely distributed in the lips, gill rakers, pharynx, buccal cavity, and also on the body surface with some variations depending on the species [9]. Interestingly, the organizations of taste buds and taste nerves along the anterior-posterior axis are mainly conserved between teleost and mammalians, though taste buds differ in areas [9,10]. Previous studies have showed that TRs are present in teleost. Teleost T1Rs show high identity to those of mammals, while some teleosts T1R2 are different, with only two or three members. In zebrafish, T1Rs consist of zfT1R1, zfT1R2a, zfT1R2b and zfT1R3, while T2Rs contain T2R1a and T2R1b [11].

In mammalian, recent studies have demonstrated that several mechanisms of innate immunity utilize components of sensory signal transduction [12,13]. Bitter and sweet taste GPCRs were considered as sentinels of defence against infection in the airway, where they played a new role in innate immunity [12,13]. TRs as detectors of pathogens. Bitter taste receptors were expressed in the upper respiratory epithelium and responded to bitter molecules released by pathogens in the mucosal environment [[14], [15], [16]]. A critical function of tuft cells in initiating mucosal responses following infection with helminths in intestinal [17]. A population of airway epithelial cells has been identified in the trachea of mice and humans with the unique morphological and transcriptional characteristics of intestinal tuft cells [[18], [19], [20]]. Another closely related cellular lineage, called solitary chemosensory cells (SCCS) [15,[21], [22], [23]], has also been identified in mouse and human nasal epithelium, but its exact relationship to tuft cells has not been determined. Both tuft cells and SCCs of the airways expressed type II taste receptors (T2Rs) in humans [15] and mice [21]. Denatonium—a potent bitter taste receptor ligand—can act on tuft cells to regulate respiration rate [24], nasal neurogenic inflammation [25]. SCCs also expressed canonical sweet taste receptors (T1R2/3), but their activation suppressed calcium flux and bitter taste receptor-induced antimicrobial responses [26]. It is suggested that SCCs, and perhaps airway tuft cells, utilize chemosensory machinery to ‘taste’ the upper respiratory tract environment and regulate innate immunity [27]. During the activation of T2R38, Nitric Oxide (NO) would be generated, and a part will diffuse into the airway surface fluid (ASL), replacing Pseudomonas aeruginosa with direct bactericidal effects [14,[28], [29], [30]]. Moreover, it has been demonstrated that T2R38 functions in airway ciliated cells as a sentinel receptor to detect bacteria and regulate innate immune responses [31]. Extra-oral TRs in the nose and paranasal sinuses serve to detect pathogens and modulate innate immune responses. Bitter taste receptors can detect bacterial byproducts, while sweet receptors were thought to release inhibition on the taste cascade when pathogens deplete glucose on the apical microenvironment. Human genetic polymorphisms in bitter taste receptors correlate with sinonasal disease and can be evaluated through oral taste testing [15]. It has been proposed that T1R2 and T1R3, along with bitter taste receptors, may play roles in gastric emptying and intestinal motility by regulating the secretion of other actors, such as hormones [32]. The roles of the sweet taste receptor beyond the oral cavity are important for providing input on the caloric and macronutrient contents of ingested food [33]. In contrast, fish gustatory tissues may be subjected to strong continuous stimulation from different aquatic pathogenic microorganisms, including bacteria, viruses and parasites. We previously found mucosa-associated lymphoid tissue in gustatory organ (buccal and pharyngeal mucosa) and strong immune response can initiate in defensing parasitic invasion [34,35]. Whether the taste receptor cells at rainbow trout mucosa are similar to tuft cells that play a role in the aspect of taste and immune functions, and whether their functions are different in different tissues remains to be further studied. Whether the association between the TRs and mucosal immunity has evolved in teleost remains to be determined.

In order to gain further insights into the role of TRs in fish mucosal immunity, we identified and analyzed the characters of three taste receptor genes in rainbow trout (Oncorhynchus mykiss): T1R1, T1R3 and T2R4. The expression of them was detected in different mucosal tissues (BM, PM, and skin) containing taste buds. Moreover, we developed three models by bath infection: bacterial, viral and parasitic pathogens, respectively. We found that expression of T1R1, T1R3 and T2R4 changed significantly in different mucosal tissues at different time points in response to three infection models, may indicate TRs are related to immunity in early vertebrates.